Methods of incorporating radiohalogens into organic molecules are known [Bolton, J. Lab. Comp. Radiopharm., 45, 485-528 (2002)]. For the case of 123I-labelled radiopharmaceuticals, Eersels et al [J. Lab. Comp. Radiopharm., 48, 241-257 (2005)] have compared the 4 principal synthetic routes:                (i) oxidative radioiodination;        (ii) nucleophilic isotopic exchange;        (iii) nucleophilic non-isotopic exchange;        (iv) electrophilic labelling.Route (iv) typically involves the use of an organometallic precursors, such as trialkyltin, trialkylsilyl or organomercury or organothallium derivative. Of these, the radioiododestannylation route was acknowledged as having become the preferred electrophilic labelling method, due to the possibility of regiospecific radioiodination at room temperature. Eersels et al concluded, however, that there was no overall preferred radioiodination method, since the choice depends on the nature of the compound to be radioiodinated.        
The use of organotin intermediates in radiopharmaceutical synthesis has been reviewed by Ali et al [Synthesis, 423-445 (1996)]. Kabalka et al have published extensively on the use of organoborane precursors to permit radioisotope and radiohalogen labelling [see eg. J. Lab. Comp. Radiopharm., 50, 446-447 and 888-894 (2007)].
Click chemistry is a growing area of biotechnology research [“Click Chemistry for Biotechnology and Materials Science”, J. Lahann (Ed), Wily, (2009)]. Hein et al have reported that 1-iodoalkynes undergo click cyclisation reactions with azides as follows [Ang. Chem. Int. Ed. Engl., 48, 8018-8021 (2009)]:

Hein is concerned with internal alkynes (ie. R is not H), and is silent on radiochemistry.
The applications of “click chemistry” in biomedical research, including radiochemistry, have been reviewed by Nwe et al [Cancer Biother. Radiopharm., 24(3), 289-302 (2009)]. As noted therein, the main interest has been in the PET radioisotope 18F (and to a lesser extent 11C), plus “click to chelate” approaches for radiometals suitable for SPECT imaging such as 99mTc or 111In. Glaser and Robins have reviewed the use of click chemistry in PET radiochemical labelling reactions, focusing on the radioisotopes 18F and 11C [J. Lab. Comp. Radiopharm., 52, 407-414 (2009)].
18F click-labelling of targeting peptides, giving products incorporating an 18F-fluoroalkyl-substituted triazole have been reported by Li et al [Bioconj. Chem., 18(6), 1987-1994 (2007)], and Hausner et al [J. Med. Chem., 51(19), 5901-5904 (2008)].
WO 2006/067376 discloses a method for labelling a vector comprising reaction of a compound of formula (I) with a compound of formula (II):
R*-L2-N3  (II)
or, a compound of formula (III) with a compound of formula (IV):
R*-L4-≡  (IV)
in the presence of a Cu(I) catalyst, wherein:
L1, L2, L3, and L4 are each Linker groups;
R* is a reporter moiety which comprises a radionuclide;
to give a conjugate of formula (V) or (VI) respectively:
R* of WO 2006/067376 is a reporter moiety which comprises a radionuclide, e.g. a positron-emitting radionuclide. Suitable positron-emitting radionuclides for this purpose are said to include 11C, 18F, 75Br, 76Br, 124I, 82Rb, 68Ga, 64Cu and 62Cu, of which 11C and 18F are preferred. Other useful radionuclides are stated to include 123I, 125I, 131I, 211At, 99mTc, and 111In.
WO 2007/148089 discloses a method for radiolabelling a vector comprising reaction of a compound of formula (I) with a compound of formula (II):
R*-L2-C≡N+—O−  (II)
or, a compound of formula (III) with a compound of formula (IV):
R*-L4-≡  (IV)
in the presence of a Cu(I) catalyst, wherein:
L1, L2, L3, and L4 are each Linker groups;
R* is a reporter moiety which comprises a radionuclide;
to give a conjugate of formula (V) or (VI) respectively:

In both WO 2006/067376 and WO 2007/148089, metallic radionuclides are stated to be suitably incorporated into a chelating agent, for example by direct incorporation by methods known to the person skilled in the art. Neither WO 2006/067376 nor WO 2007/148089 discloses any methodology specific for click radioiodination—in particular which combination of compounds of formulae (I)-(IV), together with which combinations of linker groups L1, L2, L3, L4, and which type of R* group would be suitable.
WO 2006/116629 (Siemens Medical Solutions USA, Inc.) discloses a method of preparation of a radiolabelled ligand or substrate having affinity for a target biomacromolecule, the method comprising:                (a) reacting a first compound comprising                    (i) a first molecular structure;            (ii) a leaving group;            (iii) a first functional group capable of participating in a click chemistry reaction; and optionally,            (iv) a linker between the first functional group and the molecular structure, with a radioactive reagent under conditions sufficient to displace the leaving group with a radioactive component of the radioactive reagent to form a first radioactive compound;                        (b) providing a second compound comprising                    (i) a second molecular structure;            (ii) a second complementary functional group capable of participating in a click chemistry reaction with the first functional group, wherein the second compound optionally comprises a linker between the second compound and the second functional group;                        (c) reacting the first functional group of the first radioactive compound with the complementary functional group of the second compound via a click chemistry reaction to form the radioactive ligand or substrate; and        (d) isolating the radioactive ligand or substrate.        
WO 2006/116629 teaches that the method therein is suitable for use with the radioisotopes: 124I, 18F, 11C, 13N and 15O with preferred radioisotopes being: 18F, 11C, 123I, 124I, 127I, 131I, 76Br, 64Cu, 99mTc, 90Y, 67Ga, 51Cr, 192Ir, 99Mo, 153Sm and 201Tl. WO 2006/116629 teaches that other radioisotopes that may be employed include: 72As, 74As, 75Br, 55Co, 61Cu, 67Cu, 68Ga, 68Ge, 125I, 132I, 111In, 52Mn, 203Pb and 97Ru. WO 2006/116629 does not, however, provide any specific teaching on how to apply the method to the radioiodination of biological molecules.
Qu et al [J. Med. Chem., 50(14), 3380-3387 (2007)] disclose the preparation of 125I-labelled amines:
R=MeNH—C6H4— or Me2N—C6H4—
The non-radioactive iodophenyl-triazole precursor is prepared via click cyclisation of R—≡—H and I—C6H4—N3. WO 2008/131148 is a corresponding patent application by Qu et al which discloses similar chemistry.
Dong et al [ChemBioChem, 10, 1149-1151 (2009)] disclose a method of iodinating proteins using click chemistry:

Dong et al introduced the alkyne group into the protein using the synthetic amino acid homopropargylglycine in place of the N-terminus Met residue. The protein studied is engineered to be cysteine-free, and consists of 90 amino acids. Dong et al suggest that the method could usefully be applied to radioimmunoassay using the radioisotope 125I, but no radioactive chemistry is described.
WO 2010/131745 discloses 18F-labelled phenylazides of formula shown, and their use to radiolabel alkyne-functionalised oligonucleotides via click cycloaddition, forming triazole rings:

There is therefore still a need for alternative radioiodination methods, which provide radioiodinated biological targeting molecules suitable for in vivo imaging, with the iodine label in a chemical form which is resistant to in vivo metabolic deiodination.
The Present Invention.
The present invention provides methodology for the radioiodination of biological targeting molecules (BTMs), using click radioiodination. The method has the advantage that it can be carried out under mild conditions, and is hence compatible with a range of biological molecules—potentially including such molecules where conventional direct radioiodination methods may be non-viable due to instability of the BTM under the radioiodination reaction conditions. Examples of such sensitivity includes incompatibility or instability with the oxidising conditions necessary for conventional radioiodination. The present method provides radioiodination methodology which can be carried out under non-oxidising conditions, and is hence particularly advantageous for labelling BTMs which are susceptible to oxidation.
The click cyclisation used in the present method is highly selective, hence the radioiodine labelling occurs regioselectively at the alkyne group of the functionalised BTM of Formula I. The method provides products in which the radioiodine is directly bonded to a group (Y1), which is chosen to be resistant to in vivo metabolism of the carbon-iodine bond. The radioiodinated products are thus expected to exhibit good stability with respect to metabolic deiodination in vivo, with consequent unwanted stomach and/or thyroid uptake of radioiodine. The products are therefore suitable for use as radiopharmaceuticals for in vivo imaging, which is an important advantage.
The click radioiodination methodology is readily adaptable to use with an automated synthesizer apparatus. Due to its selectivity, it can also permit BTM labelling without the need to use protecting groups. In addition, the labelling has the potential to achieve higher radiolabelling yields than conventional conjugation reactions using e.g. active ester coupling.